V O L U M E 2 9 䡠 N U M B E R 1 8 䡠 J U N E 2 0 2 0 1 1
`
`JOURNAL OF CLINICAL ONCOLOGY
`
`O R I G I N A L R E P O R T
`
`Phase I Study of Oral Azacitidine in Myelodysplastic
`Syndromes, Chronic Myelomonocytic Leukemia, and Acute
`Myeloid Leukemia
`Guillermo Garcia-Manero, Steven D. Gore, Christopher Cogle, Renee Ward, Tao Shi, Kyle J. MacBeth,
`Eric Laille, Heidi Giordano, Sarah Sakoian, Elias Jabbour, Hagop Kantarjian, and Barry Skikne
`
`A
`
`B
`
`S
`
`T
`
`R
`
`A
`
`C
`
`T
`
`Purpose
`To determine the maximum-tolerated dose (MTD), safety, pharmacokinetic and pharmacody-
`namic profiles, and clinical activity of an oral formulation of azacitidine in patients with
`myelodysplastic syndromes (MDSs), chronic myelomonocytic leukemia (CMML), or acute
`myeloid leukemia (AML).
`
`Patients and Methods
`Patients received 1 cycle of subcutaneous (SC) azacitidine (75 mg/m2) on the first 7 days of cycle
`1, followed by oral azacitidine daily (120 to 600 mg) on the first 7 days of each additional 28-day
`cycle. Pharmacokinetic and pharmacodynamic profiles were evaluated during cycles 1 and 2.
`Adverse events and hematologic responses were recorded. Cross-over to SC azacitidine was
`permitted for nonresponders who received ⱖ 6 cycles of oral azacitidine.
`
`Results
`Overall, 41 patients received SC and oral azacitidine (MDSs, n ⫽ 29; CMML, n ⫽ 4; AML, n ⫽ 8).
`Dose-limiting toxicity (grade 3/4 diarrhea) occurred at the 600-mg dose and MTD was 480 mg.
`Most common grade 3/4 adverse events were diarrhea (12.2%), nausea (7.3%), vomiting (7.3%),
`febrile neutropenia (19.5%), and fatigue (9.8%). Azacitidine exposure increased with escalating
`oral doses. Mean relative oral bioavailability ranged from 6.3% to 20%. Oral and SC azacitidine
`decreased DNA methylation in blood, with maximum effect at day 15 of each cycle. Hematologic
`responses occurred in patients with MDSs and CMML. Overall response rate (ie, complete
`remission, hematologic improvement, or RBC or platelet transfusion independence) was 35% in
`previously treated patients and 73% in previously untreated patients.
`
`Conclusion
`Oral azacitidine was bioavailable and demonstrated biologic and clinical activity in patients with
`MDSs and CMML.
`
`J Clin Oncol 29:2521-2527. © 2011 by American Society of Clinical Oncology
`
`INTRODUCTION
`
`Azacitidine is a cytidine nucleoside analog with a
`mechanism of action that involves incorporation
`into DNA and RNA.1,2 Data suggest that patients
`must be exposed to azacitidine over several treat-
`ment cycles for optimal therapeutic effect.3 The
`requirement for chronic exposure can be ex-
`plained by drug pharmacokinetics, as azacitidine
`has a short plasma half-life, and by mechanism of
`action, as induction of DNA hypomethylation
`through incorporation into DNA is cell-cycle
`dependent (S-phase restricted) and DNA rem-
`ethylation is observed by the end of each treat-
`ment cycle.4
`
`A treatment regimen facilitating chronic ad-
`ministration may help achieve optimal efficacy
`outcomes. An oral azacitidine formulation would
`improve convenience of administration and ex-
`pand the possibilities of exploring novel mainte-
`nance schedules, targeting different malignancies,
`and testing multiple combinations. A phase 0 trial
`demonstrated that a single oral azacitidine dose
`resulted in detectable levels in the blood.5
`This phase I study sought to identify the
`maximum-tolerateddose(MTD),dose-limitingtoxic-
`ities (DLTs), safety, pharmacokinetic and pharmaco-
`dynamicprofiles,andclinicalactivityoforalazacitidine
`in patients with myelodysplastic syndromes (MDSs),
`chronic myelomonocytic leukemia (CMML), or acute
`myeloid leukemia (AML).
`
`© 2011 by American Society of Clinical Oncology
`
`2521
`
`Guillermo Garcia-Manero, Elias Jabbour,
`Hagop Kantarjian, University of Texas
`MD Anderson Cancer Center, Houston,
`TX; Steven D. Gore, Sarah Sakoian, The
`Sidney Kimmel Comprehensive Cancer
`Center at Johns Hopkins, Baltimore,
`MD; Christopher Cogle, University of
`Florida, Gainesville, FL; Renee Ward,
`Tao Shi, Kyle J. MacBeth, Eric Laille,
`Heidi Giordano, Barry Skikne, Celgene,
`Summit, NJ; Barry Skikne, University of
`Kansas Medical Center, Kansas City, KS.
`
`Submitted December 21, 2010;
`accepted March 22, 2011; published
`online ahead of print at www.jco.org on
`May 16, 2011.
`
`Supported by Celgene; editorial support
`in the preparation of this manuscript
`from Excerpta Medica, sponsored by
`Celgene.
`
`Authors’ disclosures of potential con-
`flicts of interest and author contribu-
`tions are found at the end of this
`article.
`
`Clinical Trials repository link available on
`JCO.org.
`
`Corresponding author: Guillermo Garcia-
`Manero, MD, University of Texas, MD
`Anderson Cancer Center, Box 428,
`1515 Holcombe Blvd, Houston, TX
`77025; e-mail: ggarciam@mdanderson
`.org.
`
`© 2011 by American Society of Clinical
`Oncology
`
`0732-183X/11/2918-2521/$20.00
`
`DOI: 10.1200/JCO.2010.34.4226
`
`CELGENE 2084
`APOTEX v. CELGENE
`IPR2023-00512
`
`Downloaded from ascopubs.org by 72.165.200.210 on March 25, 2021 from 072.165.200.210
`Copyright © 2021 American Society of Clinical Oncology. All rights reserved.
`
`

`

`Garcia-Manero et al
`
`PATIENTS AND METHODS
`
`The trial was approved by the relevant institutional review boards and ethics
`committees. All patients gave written informed consent.
`
`Patients
`Eligible patients were ⱖ 18 years, had an Eastern Cooperative Oncology
`Group performance status score of 0 to 2, and a diagnosis of MDSs, CMML, or
`AML according to WHO classification.6,7 For patients with AML, eligibility
`was limited to those for whom standard curative measures did not exist or were
`no longer effective. Exclusion criteria included a diagnosis of acute promyelo-
`cytic leukemia, previous treatment with hypomethylating agents within 4
`weeks before cycle 1, and anticancer therapy within 21 days before the first dose
`
`Table 1. Baseline Patient Characteristics (N ⫽ 41)
`
`Parameter
`
`No. of Patients
`
`%
`
`Median age, years
`Range
`Sex
`Male
`Female
`MDSs (WHO classification)
`RA/RARS/RCMD
`RAEB-1
`RAEB-2
`MDSs-U
`CMML
`AML
`De novo
`Transformed from MDSs
`IPSS (MDSs patients)ⴱ
`Low risk
`Intermediate 1 risk
`Intermediate 2 risk
`High risk
`Not available†
`Hematology
`Median hemoglobin, g/dL
`Range
`Median white blood cell count ⫻109/L
`Range
`Median absolute neutrophil count ⫻109/L
`Range
`Median platelet count ⫻109/L
`Range
`Cytogenetics‡
`Normal chromosomal karyotype
`1 chromosomal abnormality
`2 chromosomal abnormalities
`ⱖ 3 chromosomal abnormalities
`Prior treatment with hypomethylating agent
`MDSs
`CMML
`AML
`
`70
`31-91
`
`32
`9
`29
`11
`12
`5
`1
`4
`8
`4
`4
`
`2
`12
`13
`1
`1
`
`9.3
`6.9-15.1
`2.4
`0.4-30.2
`0.8
`0.0-21.7
`54.0
`3.0-262.0
`
`17
`9
`3
`6
`16
`13
`0
`3
`
`78
`22
`71
`27
`29
`12
`2
`10
`20
`10
`10
`
`7
`41
`45
`3
`3
`
`49
`26
`9
`17
`39
`32
`0
`7
`
`Abbreviations: AML, acute myeloid leukemia; CMML, chronic myelomono-
`cytic leukemia; IPSS, International Prognostic Scoring System; MDSs, myelo-
`dysplastic syndromes; MDSs-U, MDSs unclassified; RA, refractory anemia;
`RAEB, RA with excess blasts; RARS, RA with ringed sideroblasts; RCMD,
`refractory cytopenias with multilineage dysplasia.
`ⴱIPSS score11 was available for 28 patients with MDSs.
`†Patient had a bone marrow transplantation and therefore IPSS risk was not
`considered applicable.
`‡Cytogenetic data were available for 35 patients.
`
`of study drug, or less than full recovery from any significant toxic effects of
`prior treatments.
`
`Study Design and Therapy
`This open-label, phase I, dose-escalation trial was performed in four
`participating institutions and evaluated multiple cycles of oral azacitidine
`administered daily for the first 7 days of a 28-day cycle. The objectives were to
`determine the MTD, DLTs, and the safety profile of oral azacitidine. Pharma-
`cokinetic and pharmacodynamic profiles of oral and subcutaneous (SC) azaci-
`tidine, administered on the same 7-day schedule, were also compared. A
`secondary objective was to assess the clinical activity of oral azacitidine.
`During cycle 1, patients received azacitidine 75 mg/m2 daily SC for 7 days
`of a 28-day cycle. During cycle 2 and beyond, patients received oral azacitidine
`under fasting conditions (ie, no food for 2 hours before and after dosing). The
`dose of oral azacitidine was escalated following a standard phase I 3 ⫹ 3 design.
`The starting dose was 120 mg and doses were escalated in 60 mg increments up
`to a dose of 360 mg, followed by 120 mg increments until the MTD was
`reached. Intrapatient dose escalation was permitted if the dose level to which
`the patient was escalated was associated with a DLT rate of ⱕ 33%. Treatment
`continued until disease progression, lack of activity, unacceptable toxicity, or
`patient preference.
`The MTD was defined as the highest dose at which no more than 33% of
`patients experienced a DLT. DLT was defined as: grade ⱖ 3 nausea, diarrhea,
`or vomiting despite adequate/maximal medical intervention; grade ⱖ 3 clini-
`cally significant nonhematologic toxicity unrelated to underlying disease or
`intercurrent illness; failure to recover to an absolute neutrophil count (ANC)
`of higher than 500/␮L and/or platelet count of higher than 25,000/␮L with
`hypocellular bone marrow (⬍ 5%) 42 days after starting oral azacitidine
`(patients with a baseline ANC of ⱕ 500/␮L and/or platelet count of
`ⱕ 25,000/␮L were not evaluable for neutrophil or platelet toxicity); any
`treatment-related effect resulting in missing ⱖ 3 oral azacitidine doses in the
`7-day treatment period; or any treatment-related nonhematologic toxicity
`delaying initiation of the second oral azacitidine cycle by longer than 14 days.
`Only DLTs that occurred during the first oral azacitidine cycle were considered
`in determining the MTD. Adverse events were graded using the National
`Cancer Institute Common Toxicity Criteria for Adverse Events version 3.0.
`
`Table 2. Incidence of Adverse Events According to Severity in ⱖ 20% of
`Patients Treated With Oral Azacitidine (n ⫽ 41)
`
`CTCAE Grade
`
`System Organ Class
`Preferred Term
`(MeDRA 10.1)
`
`Diarrhea
`Nausea
`Constipation
`Vomiting
`Abdominal pain
`Headache
`Fatigue
`Peripheral edema
`Fever
`Cough
`Contusion
`Dizziness
`Febrile neutropenia
`
`1
`
`2
`
`3
`
`4
`
`Total
`
`No. % No. % No. % No. % No. %
`
`10 24.4 12 29.3
`8 19.5 10 24.4
`9 22.0
`7 17.1
`4
`9.8
`6 14.6
`6 14.6
`4
`9.8
`7 17.1
`5 12.2
`6 14.6
`2
`4.9
`11 26.8
`1
`2.4
`6 14.6
`2
`4.9
`7 17.1
`1
`2.4
`9 22.0
`0
`5 12.2
`3
`0
`0
`
`7.3
`
`4
`3
`0
`3
`0
`1
`4
`0
`2
`2
`0
`0
`8
`
`9.8
`7.3
`
`7.3
`
`2.4
`9.8
`
`4.9
`4.9
`
`19.5
`
`1
`0
`0
`0
`0
`0
`0
`0
`0
`0
`0
`0
`0
`
`2.4 27 65.9
`21 51.2
`16 39.0
`13 31.7
`10 24.4
`13 31.7
`12 29.3
`12 29.3
`10 24.4
`10 24.4
`9 22.0
`8 19.5
`8 19.5
`
`NOTE. This Table includes all adverse events which started during any dosing
`cycle at which oral azacitidine was administered. Percentages are based on
`the number of patients who received at least one dose of oral azacitidine.
`Multiple reports of the same preferred term from a patient are counted only
`once, using the maximum CTCAE grade.
`Abbreviations: CTCAE, National Cancer Institute Common Toxicity Criteria
`for Adverse Events; MedDRA, Medical Dictionary for Regulatory Activities.
`
`2522
`
`© 2011 by American Society of Clinical Oncology
`
`JOURNAL OF CLINICAL ONCOLOGY
`
`Downloaded from ascopubs.org by 72.165.200.210 on March 25, 2021 from 072.165.200.210
`Copyright © 2021 American Society of Clinical Oncology. All rights reserved.
`
`

`

`Phase I Study of Oral AZA in MDSs, CMML, and AML
`
`Pharmacokinetic Analysis
`Plasma and urine pharmacokinetic evaluation of azacitidine was per-
`formed on days 1 and 7 in cycles 1 and 2. Samples were collected up to 8 hours
`after administration and analyzed using a validated high-performance liquid
`chromatography/tandem mass spectrometric method. Parameters calculated
`using noncompartmental method, included maximum observed plasma con-
`centration (Cmax), time of maximum observed plasma concentration (Tmax),
`area under the plasma concentration–time curve from zero to infinity
`(AUCinf), apparent total clearance (CL/F), relative oral bioavailability (F), and
`apparent volume of distribution (Vd/F).
`Pharmacodynamic Analysis
`DNA methylation levels were measured to determine DNA hypomethy-
`lating activity of azacitidine when administered SC or orally. Whole blood was
`collected at baseline and before drug administration on days 3, 8, 15, and 22 of
`cycle 1, and days 1, 3, 8, 15, 22, and 28 of cycle 2. Genomic DNA was purified
`from each whole blood sample using the PAXgene Blood DNA System (Qia-
`gen; Valencia, CA). DNA methylation was analyzed using the Infinium Hu-
`man Methylation27 BeadArray (Illumina; San Diego, CA). In cycle 1, DNA
`methylation data were generated from blood samples of 15 patients. For 10 of
`these patients, data were also generated in cycle 2. A methylation ratio, or beta
`
`value, for each locus per sample was calculated as methylated signal/(methyl-
`ated ⫹ unmethylated signal). Those with detection P ⱕ .05 were considered
`high-quality measures. Samples with more than 25,200 high-quality beta val-
`ues and 26,304 autosomal loci with high-quality beta values in at least half of
`the samples were used for analyses. The low-quality beta values were reim-
`puted using the pamr.knnimpute function from the R package pamr.8 Wil-
`coxon signed-rank tests were performed to identify loci with significant
`methylation differences at each post-treatment time point versus baseline;
`P ⬍ .01 was considered statistically significant. All statistical analyses were
`carried out in R (R Foundation for Statistical Computing, Vienna, Austria,
`http://www.R-project.org).
`Clinical Activity
`Data for clinical activity were evaluated using International Working
`Group (IWG) 2006 criteria, with modifications as described below, for pa-
`tients with MDSs or CMML9 and IWG 2003 criteria for patients with AML.10
`Complete remission (CR), hematologic improvement (HI), and RBC and
`platelet transfusion independence (TI) were evaluated for patients with MDSs
`or CMML. Bone marrow CR (mCR) was also evaluated but not included in the
`overall response rate. RBC transfusion dependence at baseline was defined
`as ⱖ 4 RBC units in the 56 days before cycle 1. Platelet transfusion dependence
`
`SC AZA
`
`Oral AZA
`
`8,000
`
`6,000
`
`4,000
`
`2,000
`
`0
`
`Baseline
`
`Day 3
`
`Day 8
`
`Day 15
`Day 22
`Cycle 1
`
`End
`
`Day 3
`
`End
`
`Day 8
`
`Day 15
`Day 22
`Cycle 2
`
`SC azacitidine
`cycle 1, day 15
`
`Oral azacitidine
`cycle 2, day 15
`
`63
`5,435
`
`1
`1,482
`
`8
`118
`
`No. of Hypermethylated Loci
`
`B
`
`D
`
`SC 75 mg/m2, day 1 (n = 42)
`Oral 480 mg, day 1 (n = 14)
`
`1
`
`2
`
`3
`4
`5
`Time (hours)
`
`6
`
`7
`
`8
`
`SC AZA
`
`Oral AZA
`
`900
`
`800
`
`700
`
`600
`
`500
`
`400
`
`300
`
`200
`
`100
`
`0
`
`0.2
`
`0.1
`
`0.0
`
`-0.1
`
`-0.2
`
`-0.3
`
`-0.4
`
`A
`
`Concentration (ng/mL)
`
`Azacitidine Plasma
`
`C
`
`Change Versus Baseline
`
`Average Methylation
`
`Baseline
`
`Day 3
`
`Day 8
`
`Day 15
`Day 22
`Cycle 1
`
`End
`
`Day 3
`
`End
`
`Day 8
`
`Day 15
`Day 22
`Cycle 2
`
`(B)
`(A) Mean azacitidine (AZA) plasma concentration versus time profiles following single subcutaneous (SC) or oral administration (linear scale).
`Fig 1.
`Pharmacodynamics as measured by plotting the numbers of highly methylated loci (beta ⱖ 0.7; ⫾ 95% CI) for 10 patients with DNA methylation data in cycles 1 and
`2 (gold lines represent individual patients, blue line represents the average). (C) Change in methylation level during treatment with SC or oral AZA for 5,232 loci highly
`methylated at baseline (blue box represents the 25th to 75th percentile, horizontal band represents the median, vertical line with bars represents minimum and
`maximum values). (D) Number of significantly differentially methylated loci on day 15 of cycle 1 (SC azacitidine) and on day 15 of cycle 2 (oral azacitidine). Upward arrows
`denote hypermethylated loci and downward arrows denote hypomethylated loci.
`
`www.jco.org
`
`© 2011 by American Society of Clinical Oncology
`
`2523
`
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`

`

`Garcia-Manero et al
`
`at baseline was defined as ⱖ 2 platelet transfusions in the 56 days before cycle 1
`(modification to IWG 2006 criteria). RBC and platelet TI were defined as no
`transfusions in any 56 consecutive-day period on treatment. Patients who
`achieved ⱖ 50% reduction in platelet transfusion requirement, but not platelet
`TI, in any 56 consecutive-day period on treatment were counted as having
`achieved HI platelet (HI-P; modification to IWG 2006 criteria). Patients RBC
`transfusion dependent at baseline achieving a ⱖ 50% reduction in RBC trans-
`fusion requirement in any 56-consecutive day period and patients not RBC
`transfusion dependent at baseline, but who achieved a 1.5 g/dL increase in
`hemoglobin in any 56-consecutive day period on treatment were considered
`to have achieved HI erythroid (HI-E; modification to IWG 2006 criteria). All
`patients who received ⱖ 1 cycle of oral azacitidine were included in the
`response analysis. The cutoff date for data in this article was August 19, 2010.
`
`RESULTS
`
`Patient Characteristics
`Forty-five patients were treated on a 7-day once-daily schedule.
`Four patients received the first cycle of SC azacitidine only; three
`discontinued due to progressive disease (including one death), and
`one withdrew consent. Baseline characteristics for the remaining 41
`patients who received oral azacitidine are presented in Table 1.11
`Cytogenetic data were available at baseline for 35 of 41 patients treated
`with oral azacitidine; nearly half of the patients had normal karyotype,
`approximately 25% had a single abnormality, and nearly 20% had a
`complex karyotype (ⱖ 3 chromosomal abnormalities). Overall, 16
`(39%) of 41 patients had received prior hypomethylating therapy.
`
`Dose Escalation of Oral Azacitidine
`No DLTs were observed at dose levels up to 480 mg. DLT was
`observed at the 600 mg dose, with two (66.7%) of three patients
`experiencing severe diarrhea, despite adequate medical intervention
`(grade 3 in one patient and grade 4 in the other). Per protocol, the
`MTD was exceeded and the previous dose level of 480 mg was deter-
`mined to be the MTD.
`
`Safety Profile
`Table 2 shows the incidence of AEs (any grade) that oc-
`curred in ⱖ 20% of patients treated with oral azacitidine. The most
`
`frequently observed AEs were gastrointestinal disorders, headache,
`fatigue, and peripheral edema. Other commonly occurring AEs
`included fever, cough, contusion, dizziness, and febrile neutrope-
`nia. Grade 3/4 nausea and grade 3/4 vomiting were each observed
`in 7% of patients. Grade 3 fatigue was observed in 10% of patients.
`Diarrhea occurred at grade 3 severity in 10% of patients and grade
`4 severity in 2%. Grade 3 febrile neutropenia was observed in eight
`patients (20%), with four of those having an ANC of ⱕ 500/␮L
`at baseline.
`Of the 41 patients who received oral azacitidine, 33 terminated
`from the study as of the date of data analysis, with 17 discontinuing
`before completing 6 cycles of oral therapy. Reasons for discontinua-
`tion included disease progression/treatment failure (n ⫽ 10), investi-
`gator decision primarily due to absence of observed benefit/response
`(n ⫽ 15), withdrawal of consent (n ⫽ 4), AEs (n ⫽ 3), and decision to
`pursue hematopoietic stem-cell transplantation (n ⫽ 1). There were
`three deaths within 28 days of last dose of study drug due to multiple
`organ failure (n ⫽ 1), gastrointestinal hemorrhage (n ⫽ 1), and
`pneumonia plus urinary tract infection (n ⫽ 1). No deaths were
`attributed to study drug. Eight patients remained on the study at the
`time of data analysis, having each received between 14 and 32 treat-
`ment cycles.
`
`Pharmacokinetic Characteristics of Azacitidine
`High interpatient variability was noted for all pharmacokinetic
`parameters. Azacitidine was rapidly absorbed after SC (n ⫽ 42) and
`oral (n ⫽ 36) administration, reaching Cmax within 0.5 hours (range,
`0.2 to 1.1 hours) and 1.0 hours (range, 0.3 to 3.6 hours) postdose,
`respectively. Concentration versus time profiles decreased in a pseu-
`dobiphasic manner (Fig 1A). The mean elimination half-life was 1.6 ⫾
`0.7 hours for SC and 0.62 ⫾ 0.25 hours for oral azacitidine. Exposure
`after single oral administration generally increased with dose (Table
`3). For the seven oral dose levels, the mean relative azacitidine oral
`bioavailability (F) ranged from 6.3% to 20%. The MTD had a mean
`relative bioavailability of 13% ⫾ 9%. CL/F exceeded hepatic blood
`flow, indicating extrahepatic metabolism, and Vd/F was greater than
`total body water, suggesting extensive tissue distribution. The amount
`of azacitidine recovered in urine relative to dose was small (⬍ 2%) for
`
`Table 3. Day 1 Plasma Pharmacokinetics Parameters After Single Subcutaneous or Oral Azacitidine Administration
`
`AUCinf
`(ng ⫻ h/mL)
`Tmax (h)
`Cmax (ng/mL)
`CL/F (L/h)
`Mean SD %CV Mean
`SD %CV Mean SD %CV Median Range Mean
`
`No. of
`Patients
`
`F (%)
`
`Vd/F (L)
`SD %CV Mean SD
`
`Relative Oral
`Bioavailability
`
`42
`
`4
`3
`3
`5
`5
`14
`2
`
`1,020 440
`
`43ⴱ
`
`175
`
`128
`
`73ⴱ
`
`650
`
`250
`
`39
`
`0.50
`
`0.2-1.1
`
`410
`
`410 101ⴱ NA
`
`43
`62
`64
`112
`463 221
`282
`88
`311 141
`362 253
`502 100
`
`70
`58
`48
`31
`45
`70
`20
`
`4,100 4,860 118
`2,330 1,890
`81
`598
`258
`43
`1,180
`487
`41
`1,360
`573
`42
`2,140 1,620
`76
`1,220
`244
`20
`
`38
`72
`215
`144
`195
`211
`253
`
`24
`36
`102
`13
`79
`140
`29
`
`64
`50
`47
`9.2
`40
`66
`12
`
`1.48
`1.50
`1.00
`1.48
`1.00
`1.00
`1.50
`
`1.0-2.0 2,930 3,810 130
`1.0-1.5 1,700 1,580
`93
`1.0-1.5
`814
`421
`52
`1.0-2.0 1,090
`626
`57
`0.5-3.6
`947
`251
`27
`0.3-2.5 2,010 1,910
`95
`1.0-2.0 1,580 1,410
`89
`
`8.1
`6.3
`20.0
`11.5
`12.8
`12.8
`14.9
`
`5.6
`2.3
`9.6
`2.6
`2.4
`9.4
`0.8
`
`69
`37
`48
`23
`19
`74†
`5
`
`Dose
`
`Subcutaneous,
`75 mg/m2
`Oral, mg
`120
`180
`240
`300
`360
`480
`600
`
`Abbreviations: AUCinf, area under the plasma concentration–time curve from time zero to infinity; CL/F, apparent total clearance; Cmax, maximum observed plasma
`concentration; F, relative oral bioavailability; NA, not applicable; Tmax, time of maximum observed plasma concentration; Vd/F, apparent volume of distribution.
`ⴱn ⫽ 40.
`†n ⫽ 13.
`
`2524
`
`© 2011 by American Society of Clinical Oncology
`
`JOURNAL OF CLINICAL ONCOLOGY
`
`Downloaded from ascopubs.org by 72.165.200.210 on March 25, 2021 from 072.165.200.210
`Copyright © 2021 American Society of Clinical Oncology. All rights reserved.
`
`

`

`Phase I Study of Oral AZA in MDSs, CMML, and AML
`
`SC and oral administration, suggesting that nonrenal elimination
`is the predominant pathway for clearance. Results after multiple
`doses were similar to those obtained after a single dose for both
`administration routes (data not shown). There was no evidence of
`azacitidine accumulation.
`
`Pharmacodynamics of Azacitidine: Effect on
`DNA Methylation
`DNA methylation was evaluated during cycles 1 and 2 in 10
`patients treated with oral azacitidine. The numbers of highly methyl-
`ated loci were calculated at each time point by averaging across pa-
`tients the number of loci with methylation ratios ⱖ 0.7 (Fig 1B). These
`numbers decreased after SC and oral administration, with maximal
`effects at day 15 of each cycle. The reduction in levels of highly meth-
`ylated loci was not maintained throughout the entire cycle and re-
`turned to near-baseline levels by the end of each cycle. SC azacitidine
`decreased a greater number of loci in comparison to oral azacitidine.
`The changes in methylation level from baseline across patients for the
`5,232 highly methylated loci (average methylation ratio at baseline
`ⱖ 0.7) are represented as box plots (Fig 1C). As with the analysis of
`total numbers of highly methylated loci, the median DNA methyl-
`
`ation of these loci was reduced by 0.115 on day 15 of cycle 1 (SC
`azacitidine) and 0.055 on day 15 of cycle 2 (oral azacitidine), and
`returned to baseline levels at the end of each cycle.
`Differentially methylated loci at each post-treatment time point
`compared with baseline were identified in cycles 1 and 2, with the
`maximum number observed on day 15 of each cycle; 6,981 loci were
`differentially methylated (6,917 hypomethylated) on day 15 of cycle 1
`(SC azacitidine) and 1,609 loci were differentially methylated (1,600
`hypomethylated) on day 15 of cycle 2 (oral azacitidine; P ⬍ .01). In
`total, 1,482 loci were significantly hypomethylated by both SC and oral
`azacitidine (Fig 1D), representing 92.6% of all loci significantly hy-
`pomethylated by oral azacitidine treatment. These data demonstrate
`comparable biologic activity with SC and oral azacitidine, albeit to a
`lesser extent with oral azacitidine.
`
`Clinical Activity of Oral Azacitidine
`The median number of oral azacitidine cycles administered to
`patients with MDSs, CMML, and AML was 6 (range, 1 to 32⫹), 12.5⫹
`(range, 3 to 28⫹), and 4.5 (range, 1 to 15), respectively. Treatment
`duration is summarized in Figure 2. The number of patients from the
`MDSs, CMML, and AML groups who remained on the study at the
`
`MDSs-U
`RCMD
`RAEB-1
`RAEB-1
`RCMD
`RA
`RAEB-2
`RA
`RCMD
`RAEB-1
`RAEB-1
`RAEB-2
`RCMD
`RAEB-1
`RA
`RCMD
`RAEB-2
`RAEB-2
`RAEB-1
`RAEB-1
`RA
`RAEB-1
`RAEB-1
`RCMD
`RCMD
`RAEB-1
`RAEB-1
`RAEB-1
`RAEB-2
`
`CMML
`CMML
`CMML
`CMML
`
`AML
`AML
`AML
`AML
`AML
`AML
`AML
`AML
`
`Fig 2. Treatment duration for the 41 pa-
`tients treated with oral azacitidine (AZA).
`AML, acute myeloid leukemia; CMML,
`chronic myelomonocytic leukemia; MDSs-U,
`myelodysplastic syndromes-unclassified; RA,
`refractory anemia; RCMD, refractory cyto-
`penias with multilineage dysplasia; RAEB,
`RA with excess blasts; SC, subcutaneous.
`
`Initial SC cycle
`Oral cycles, active
`Oral cycles, discontinued
`Crossover SC cycles, discontinued
`
`0
`
`5
`
`10
`
`15
`20
`No. of Cycles Initiated
`
`25
`
`30
`
`35
`
`www.jco.org
`
`© 2011 by American Society of Clinical Oncology
`
`2525
`
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`
`

`

`Garcia-Manero et al
`
`Table 4. Response in Myelodysplastic Syndromes and Chronic Myelomonocytic Leukemia Patients
`
`Previously Treated Patientsa
`
`First-Line Treatment
`
`Response
`
`Responders
`
`Evaluable Patients
`
`Overall responseb
`CRd
`Any HIe
`HI-E
`HI-N
`HI-P
`
`TI
`
`Red blood cell
`Platelet
`mCRe,f
`
`6
`0
`6
`3
`0
`5
`0
`0
`0
`6
`
`17
`17
`16
`10
`10
`14
`5
`3
`4
`9
`
`%
`
`35
`0
`38
`30
`0
`36
`0
`0
`0
`67
`
`Responders
`
`Evaluable Patients
`
`11
`6
`5
`2
`2
`2
`1
`1
`0
`2
`
`15
`15
`9
`4
`7
`6
`3
`3
`
`6
`
`%
`
`73
`40
`56
`50
`29
`33
`33
`33
`
`33
`
`Duration of
`Response:Range
`(days)
`
`30-483c
`30-152
`56-483c
`56-483c
`82-321c
`58-351c
`76
`76
`NA
`63-422g
`
`NOTE. At any cycle of azacitidine, International Working Group 2006 criteria were used with modifications as described in the Patients and Methods section.
`Abbreviations: CR, complete remission; E, erythroid; HI, hematologic improvement; mCR, bone marrow complete remission; N, neutrophil; NA, not applicable; P,
`platelet; TI, transfusion independence.
`aIncludes erythropoiesis-stimulating agents, chemotherapy, hypomethylating agents, and investigational and/or other agents.
`bOverall response rate does not include patients achieving mCR only.
`cOne or more responses, including that at upper limit of range, are ongoing. Data were censored as of last visit entered into the clinical database.
`dPatients achieving CR were not included in any other categories.
`eOne patient with mCR in the previously treated group also achieved HI (both HI-E and HI-P). Two patients with mCR in the first-line treatment group also achieved
`HI (one patient with HI-P and one patient with both HI-E and HI-N). These patients have been included in both the mCR and HI categories.
`fIn the eight patients who achieved mCR, the response began in cycle 1 of subcutaneous (SC) dosing (n ⫽ 4) or very early in cycle 2 of oral dosing (n ⫽ 4). Therefore,
`the contribution of a single SC azacitidine cycle to the induction of these responses is likely relevant.
`gBone marrow aspirates were not required after 6 cycles of oral azacitidine treatment, therefore follow-up data were not available to confirm upper limit of duration.
`Data were censored as of last visit entered into the clinical database.
`
`time of the analysis was 6, 2, and 0, respectively. Response and dura-
`tion of response data are summarized in Table 4. In the 17 previously
`treated patients with MDSs and CMML, the overall response rate was
`35%,withoutincludingpatientswhoonlyachievedmCR;ifthosepatients
`were included the response rate would be 65%. In the 15 patients with
`MDSs and CMML receiving first-line treatment, the overall response rate
`was 73% and in this group no patients achieved mCR only. Longest
`duration of response to date was 483 days overall. In one patient who
`achievedaCR,theresponsebeganbeforeoraldosingandendedincycle2,
`thus was likely attributable to the single cycle of SC azacitidine.
`No responses were observed in patients with AML. Two patients
`with AML (25%) had stable disease for 14 and 15 cycles, respectively,
`and five patients with AML (63%) received ⱖ 4 oral azacitidine cycles.
`
`DISCUSSION
`
`An oral azacitidine formulation may bring advantages for patients
`(ease of administration), society (health care cost implications), and
`disease treatment (extended administration), provided that clinical
`activity and safety are similar to SC/intravenous azacitidine. This
`phase I trial demonstrated that oral azacitidine is associated with
`minimal adverse effects at doses lower than 600 mg. The MTD was 480
`mg on a 7-day of 28 days treatment schedule. The 600 mg dose was
`associated with early onset of severe diarrhea in two of three patients.
`Diarrhea in patients taking oral azacitidine doses lower than 600 mg
`was self-limiting and manageable by treatment and/or prophylaxis
`with antidiarrheal agents and/or dose reduction. Azacitidine, along
`with one or more ingredients used in its formulation, may contribute
`to the diarrhea observed because it was a common adverse event at
`most dose levels tested. Gastrointestinal disturbances may have been
`exacerbated by the requirement to ingest oral azacitidine in a fasting
`state. Whether oral azacitidine administration with food can reduce gas-
`
`trointestinal toxicity will be evaluated in ongoing studies. Grade 3 and 4
`AEs consisted primarily of febrile neutropenia, gastrointestinal distur-
`bances, and fatigue. Of the eight patients who experienced grade 3 febrile
`neutropenia, four entered the study with a baseline ANC of ⱕ 500/␮L.
`After oral administration, maximum azacitidine plasma concen-
`trations were achieved rapidly (within 1 hour), suggesting that absorp-
`tion occurs from the proximal gastrointestinal tract. Azacitidine
`exposure increased with increasing oral doses, and the mean relative oral
`bioavailability ranged from 6.3% to 20%. After multiple doses, there was
`no evidence of azacitidine accumulation, and no apparent decline in
`absorption was seen between days 1 and 7. Azacitidine clearance was
`hepatic and extrahepatic, with little evidence of renal clearance.
`Kinetics of the change in DNA methylation levels after SC and
`oral azacitidine were similar, with maximum hypomethylation
`achieved on day 15, and methylation levels returned to near-baseline
`values by the end of each cycle. This pattern has been observed in other
`azacitidine studies.3,12 At the dosing schedule employed in this study,
`oral azacitidine affected fewer loci than SC azacitidine; however, 1,482
`loci were identified as commonly hypomethylated by both azaciti-
`dine formulations.
`Significant responses were observed in patients with MDSs and
`CMML, indicating that oral azacitidine has clinical activity in these
`settings. Although all patients received an initial cycle of SC azaciti-
`dine, which may have contributed to the clinical activity observed, it
`has been reported that only half of the total hematologic responses to
`SC azacitidine manifest within 2 cycles.13 Continued treatment with
`oral azacitidine following the single cycle of SC azacitidine is therefore
`likely to be associated with the development and/or maintenance of
`clinical responses observed in this study.
`Results from a study investigating alternative SC azacitidine dos-
`ing schedules in lower-risk patients with MDSs suggested that for all
`
`2526
`
`© 2011 by American Society of Clinical Oncology
`
`JOURNAL OF CLINICAL ONCOLOGY
`
`Downloaded from ascopubs.org by 72.165.200.210 on March 25, 2021 from 072.165.200.210
`Copyright © 2021 American Society of Clinical Oncology. All rights reserved.
`
`

`

`Phase I Study of Oral AZA in MDSs, CMML, and AML
`
`dosing regimens tested, continued azacitidine treatment may be ben-
`eficial.14 The short plasma half-life of azacitidine, S-phase restricted
`incorporation into DNA, and rapid remethylation of DNA, are con-
`tributing factors to the importance of chronic exposure to the drug. It
`is therefore likely that extended schedules of oral administration will
`positively affect clinical activity of azacitidine. A follow-up trial has
`been initiated to investigate the efficacy of such extended schedules.
`In conclusion, the MTD for oral azacitidine administered daily
`for 7 days of a 28-day cycle was determined to be 480 mg, and oral
`azacitidine is bioavailable and biologically active. Clinical responses
`were reported in 35% of previously treated patients with MDSs and
`CMML, and in 73% of patients who received oral azacitidine as first-
`line therapy. Lower drug exposure